Molecular cloning and expression of a Toll receptor gene homologue from the silkworm, Bombyx mori

Abstract

A novel Toll receptor gene was cloned from the bacterial artificial chromosomal (BAC) library of the silkworm, Bombyx mori, using primers for polymerase chain reactions (PCR), synthesized based on an expressed-sequence tag (EST). The open reading frame of this gene was intronless. The deduced amino acid sequence including a putative signal peptide from the nucleotide sequence of the B. mori Toll (BmToll) gene shows 53.7%, 49.0%, 42.3% and 39.2% similarity to those of Toll-7, 18 wheeler (18w), Toll-6 and Toll-8 from Drosophila melanogaster. BmToll, however, does not show marked similarity to Toll, Toll-3, Toll-4 or Toll-5. Phylogenetic insect Toll family analysis shows that BmToll is strongly related to Toll-7 and 18w. An analysis of the tissue-specific expression of the BmToll gene by reverse transcription PCR (RT-PCR) showed that the BmToll gene is constitutively expressed in the fat body but not in the Malpighian tubule, silk gland, midgut or hemocyte. Injecting lipopolysaccharide (LPS) into silkworm hemocoel strongly suppressed BmToll gene expression. The time course showed that BmToll gene expression is totally suppressed within 2 h and gene transcripts appeared again 12 h after LPS injection, suggesting strong downregulation of the BmToll gene expression by LPS.